Project Summary Globally, chronic disorders including cardiovascular disease, diabetes, cancer, and chronic respiratory disorders represent one of the largest causes of death in industrialized societies. Besides life-threatening disease, chronic pain currently inflicts millions of American adults and contributes to billions ever year in medical costs. While complex molecular factors underlie these heterogeneous pathologies, a unifying feature of numerous chronic disorders is non-resolved inflammation. Thus, new anti-inflammatory targets are needed to combat the burden of chronic inflammatory disease. Macrophages accumulate at inflammatory sites to produce lipid and protein inflammatory signaling molecules that can cause profound changes in physiology including sensitization of peripheral sensory neurons to promote pathogenesis of chronic pain. We previously discovered that diacylglycerol lipase-beta (DAGLB) regulates an endocannabinoid-eicosanoid lipid-signaling network critical for activation of proinflammatory responses in macrophages. Recent preliminary data further support DAGLB-regulated lipid pathways as a safe and effective point of intervention in mouse models of inflammatory and neuropathic pain that lack gastrointestinal and overt behavioral side effects. Our proposed studies build on published data from our group as well as others that point to DAGLB-regulated pathways in macrophages as a novel anti-inflammatory target for treating chronic inflammation and pain. Here, we plan to test our central hypothesis that DAGLB-inactivation produces anti-inflammatory effects via autocrine lipid signaling pathways in macrophages to reduce local inflammatory responses in vivo. We propose a highly innovative approach that leverages chemical proteomics, mass spectrometry metabolomics, mouse pain models, and novel small molecule probes that selectivity detect and inactivate DAGLB in vivo. The impact of our proposed studies include 1) molecular elucidation of crosstalk between lipid and kinase signaling pathways that explain the anti-inflammatory effects of DAGLB inhibitors, 2) insights into the translational potential of endocannabinoid biosynthetic pathways in human innate immune biology, and 3) creation of new drug delivery strategies for targeting macrophage signaling for treating pathological pain. Our long-term goals are to identify new anti-inflammatory mechanisms and drug delivery strategies for developing targeted anti- inflammatory drugs suitable for treating chronic disease.